Molecular Biology

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    Protocols in Current Issue
    Enterovirus Competition Assay to Assess Replication Fitness
    Authors:  Valeria Lulla and Andrew E. Firth, date: 05/20/2019, view: 155, Q&A: 0
    In virology the difference between the fitness of two viruses can be determined by using various methods, such as virus titer, growth curve analysis, measurement of virus infectivity, analysis of produced RNA copies and viral protein production. ...
    Purification and Proteomic Analysis of Alphavirus Particles from Sindbis Virus Grown in Mammalian and Insect Cells
    Authors:  Raquel Hernandez, Trevor Glaros, Gabrielle Rizzo and Davis F. Ferreira, date: 05/20/2019, view: 153, Q&A: 0
    Current mass spectrometry (MS) methods and new instrumentation now allow for more accurate identification of proteins in low abundance than previous protein fractionation and identification methods. It was of interest if this method could serve to ...
    Enterovirus Competition Assay to Assess Replication Fitness
    Authors:  Valeria Lulla and Andrew E. Firth, date: 05/20/2019, view: 155, Q&A: 0
    [Abstract] In virology the difference between the fitness of two viruses can be determined by using various methods, such as virus titer, growth curve analysis, measurement of virus infectivity, analysis of produced RNA copies and viral protein production. However, for closely performing viruses, it is often very hard to distinguish the differences. In vitro ...
    Purification and Proteomic Analysis of Alphavirus Particles from Sindbis Virus Grown in Mammalian and Insect Cells
    Authors:  Raquel Hernandez, Trevor Glaros, Gabrielle Rizzo and Davis F. Ferreira, date: 05/20/2019, view: 153, Q&A: 0
    [Abstract] Current mass spectrometry (MS) methods and new instrumentation now allow for more accurate identification of proteins in low abundance than previous protein fractionation and identification methods. It was of interest if this method could serve to define the virus proteome of a membrane-containing virus. To evaluate the efficacy of mass spec to ...
    Improved HTGTS for CRISPR/Cas9 Off-target Detection
    Authors:  Jianhang Yin, Mengzhu Liu, Yang Liu and Jiazhi Hu, date: 05/05/2019, view: 270, Q&A: 0
    [Abstract] Precise genome editing is essential for scientific research and clinical application. At present, linear amplification-mediated high-throughput genome-wide translocation sequencing (LAM-HTGTS) is one of most effective methods to evaluate the off-target activity of CRISPR-Cas9, which is based on chromosomal translocation and employs a “bait” DNA ...
    Unbiased Screening of Activated Receptor Tyrosine Kinases (RTKs) in Tumor Extracts Using a Mouse Phospho-RTK Array Kit
    Authors:  Julian Naipauer, Lucas E. Cavallin and Enrique A. Mesri, date: 04/20/2019, view: 384, Q&A: 0
    [Abstract] Kaposi’s sarcoma (KS) herpesvirus (KSHV) is a virus that causes KS, an angiogenic AIDS-associated spindle-cell neoplasm, by activating host oncogenic signaling cascades through autocrine and paracrine mechanisms. Many host signaling cascades co-opted by KSHV including PI3K/AKT/mTORC, NFkB and Notch are critical for cell-specific mechanisms of ...
    Northern Blot with IR Fluorescent Probes: Strategies for Probe Preparation
    Authors:  Christopher Fields, Peike Sheng, Bret Miller, Tianqi Wei and Mingyi Xie, date: 04/20/2019, view: 447, Q&A: 0
    [Abstract] Northern blot is a molecular biology technique that can detect, quantify, and determine the molecular weight of RNA. Recently, we published a protocol utilizing near-infrared (IR) fluorescent probes in Northern blot (irNorthern). Our method is as sensitive as other non-radioactive methods but is more straightforward and versatile. Additionally, we ...
    Identification of RNase-sensitive LINE-1 Ribonucleoprotein Interactions by Differential Affinity Immobilization
    Authors:  Hua Jiang, Martin S. Taylor, Kelly R. Molloy, Ilya Altukhov and John LaCava, date: 04/05/2019, view: 626, Q&A: 0
    [Abstract] Long Interspersed Nuclear Element-1 (LINE-1, L1) constitutes a family of autonomous, self-replicating genetic elements known as retrotransposons. Although most are inactive, copious L1 sequences populate the human genome. L1s proliferate in a ‘copy-and-paste’ fashion through an RNA intermediate; a full-length L1 transcript is ~6,000 nucleotides ...
    Multiple Modification of Chromosomal Loci Using URA5.3 Selection Marker in the Unicellular Red Alga Cyanidioschyzon merolae
    Authors:  Tokiaki Takemura, Sousuke Imamura, Yuki Kobayashi and Kan Tanaka, date: 04/05/2019, view: 381, Q&A: 0
    [Abstract] The unicellular red alga Cyanidioschyzon merolae has been used as a eukaryotic photosynthetic model for various basic and applied studies. Although the nuclear genome of C. merolae can be modified by homologous recombination with exogenously introduced DNA, it has been difficult to modify multiple chromosome loci within the same ...
    Quantification of Hepatitis B Virus Covalently Closed Circular DNA in Infected Cell Culture Models by Quantitative PCR
    Authors:  Bingqian Qu and Stephan Urban, date: 04/05/2019, view: 494, Q&A: 0
    [Abstract] Persistence of the human hepatitis B virus (HBV) requires the maintenance of covalently closed circular (ccc)DNA, the episomal genome reservoir in nuclei of infected hepatocytes. cccDNA elimination is a major aim in future curative therapies currently under development. In cell culture based in vitro studies, both hybridization- and ...
    Delivering "Chromatic Bacteria" Fluorescent Protein Tags to Proteobacteria Using Conjugation
    Authors:  Rudolf O Schlechter and Mitja NP Remus-Emsermann, date: 04/05/2019, view: 464, Q&A: 0
    [Abstract] Recently, we published a large and versatile set of plasmids, the chromatic bacteria toolbox, to deliver eight different fluorescent protein genes and four combinations of antibiotic resistance genes to Gram-negative bacteria. Fluorescent tags are important tools for single-cell microbiology, synthetic community studies, biofilm, and host-microbe ...
    Preparation of RNA 3’ End Sequencing Libraries of Total and 4-thiouracil Labeled RNA for Simultaneous Measurement of Transcription, RNA Synthesis and Decay in S. cerevisiae
    Authors:  Manfred Schmid, Agnieszka Tudek and Torben Heick Jensen, date: 03/20/2019, view: 562, Q&A: 0
    [Abstract] Cellular RNA levels are determined by the rates of RNA transcription from the gene template and subsequent RNA stability. Knowledge about both transcription and RNA decay is, therefore, necessary to interpret RNA levels and gene expression, especially during cellular processes where these parameters change. Numerous experimental strategies have ...



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